Sapogenin oxidation products and process for preparing the same



Patented Sept. 3, 1946 v SAPOGENIN OXIDATION PRODUCTS AND ,1 srA'rEs PATENT OFFICE PROCESS FOR PREPARING THE SAME Russell Earl'xMarker, State College, Pa., assignor 1' to Parke, Davis & Company, Detroit, Mich., a

5 corporation of Michigan" No'Di'awing. Original application August 3, 1940,

Divided and this applicaa v tion November 22,1943, Serial No. 511,340

Serial No. 351,147.

.f The invention relates to 'sapogeninoxidation product's; andmore particularly tocompounds which may be described v v as Cz-keto acids, having in ring D the following formula,

CHa (I? where R is a member of the class consisting of OH and groups hydrolyzable to -OH.

This application is a division of my copending application, Serial No. 351,147, filed August 3, 19%10.

The steroidal sapogenins havaingeneral, the formula C27H42-4O3-5, of which the portion CaHmOz is known to be present as a side chain attached to ring D of the steroid skeleton.

The following sapogenins differ from one another rnainly in regardto the connections between rings A and B, 'the degree ,of. saturation of the ring system, and the number of subisola s, (01. 260-3971) 3 Dlosg e nln I Sarsasapogenln Recently it has been found (Marker and Ruhrmann, J. Am. Chem.Soc. 61, 846, 1516, 2724, 3479, (1939); (i2, 647, 896, 1162) that the side chain of the naturally occurring sapogenins exists in 1 two modifications. Sarsasapogeninand neotigo genin contain one type of side chain characterited by the fact that a sapogenin of this type is readily reduced according to the Clemmensen procedure using alcoholic hydrochloric acid, and amalgamated zinc (Marker and Rohrmann, J. Am. Chem. Soc. 61, 846 (1939)) to give tetrahydro derivatives, that is to say, new sapogenin derivatives in which the side chain has 4 more hydrogen atoms than in the sapogenins themselves. Tigogenin, gitogenin, digitogenin, chlorogenin and diosgenin behave differently from sarsasapogenin and neotigogenin in that they are not reduced by the Clemmensen procedure, that is to say, they are recovered unchanged after boiling with alcoholic hydrochloric acid and Y amalgamated zinc. Isosarsasapogenin is converted under the conditions of the Clemmensen reduc tion to the same tetrahydrosarsasapogenin that sarsasapogenin itself yields. These facts, and others which are cited in the references listed, appear to me to be best explained by assuming that thetwo types of steroidal sapogenin side chains differ in regard to optical isomerism about Accordingly, when it is necessary to distinguish between isomers about C22 I represent the :two types. of side chains byformulae of the following type: 1 I

CH: CH; CH;

E /CH1 1 C CH-CHa V CH2Ca Sarsasapogenin (Sarsasapogenin type" side chain) CH: CHI CH:

' 1E /CHr-CE: C\ CH-CH;

L O-Ca H0 /Y Isosarsasapogenin (Tigogenin type side chain) See especially Marker and Rohrmann, J. Am. Chem. Soc. 62, 896 (1940) Ordinarily where isomerism about C22 is not of importance, the formula for the sarsasapogenin type of side chain will be used for configurations both of sarsasapogenin and tigogenin type.

These two types of sapogenin side chains appear to be subject to an equilibrium, the velocity of attainment of which is catalyzed by acidic reagents. The equilibrium is influenced by the configuration of the hydrogen atom at C5, and it appears that the following rule holds true. For compounds of the allo series (cholestane type) the tigogenin type of side chain is the more stable, while for compounds of the regular series (coprostane type) at C5 the sarsasapogenin side chain is the more stable.

For the purpose of greater clarification, the Marker-Rohrmann formulation of the side chain of the steroidal sapogenins will be used in describing the invention. It is to be understood, however, that the processes and products of the present invention may be obtained by the use of the methods herein to be described and these processes and products are claimed without any implications that the reactions and structures involved will ultimately be proved to be as represented herein.

I have found that the steroidal sapogenins and certain derivatives thereof may be oxidized to give new sapogenin oxidation products which are valuable as intermediates for the preparation of steroidal hormones.

In practicing my invention I proceed as follows: The sapogenin or the derivative thereof which is to be oxidized is first treated with reagents to protect nuclear hydroxyl groups and/or double bonds. For instance, nuclear hydroxyl groups are treated with acylating, aralkylating or halogenating agents to form the corresponding -O-acyl, -O-aralkyl or halogen derivative. Nuclear double bonds are protected by addition of hydrogen halide or halogen. Then the nuclearly protected sapogenin or derivative thereof is oxidized, preferably with a reagent selected from the group consisting of compounds of hexavalent chromium and compounds of heptavalent manganese. The reaction mixture is separated into alkali-soluble and alkali-insoluble fractions, and each of these fractions is further separated into its components by crystallization, partition between immiscible solvents, conversion into insoluble derivatives, high vacuum distillation, chromatographic adsorption or other methods adapted to the properties of the substances to be isolated.

My invention also comprehends certain new classes of steroidal sapogenin oxidation products. One of these classes of compounds may be represented by the following formula CH: O CHI 4} ll nA CH:

Y A B where the symbol nA represents 11, carbon-t0- carbon double bonds in the A-B portion of the steroid nucleus, n having one of the values 0 and 1, Y1, Y2 and Y: are substituents attached-to methylene carbon atoms in the AB portion of the steroid nucleus said substituents being selected from the class consisting of groups hydrolyzable to and groups hydrolyzable to OH O-acyl O-aralkyl include H halogen and Groups hydrolyzable to-OH include O-alkyl, --NH2, and halogen.

My invention may be illustrated by the following partial formulae in which only transformations of the side chain attached to ring D are 3 shown;

C CH1". CH: 6H CH:CH: H CHI CH-CHa D B-CH|CHa HCHaOE o-caz cat.

hydrog Sapogenln Dlhydrosapogenln C 01-0; 25260 0. 12s c.

CH: CH: CHI n (EH-fir-CHICILCHCOOH CH CH: 25- 0 /K'JH CH: 70C. 0 EH-CHaCHaJJHOOOB Sapogenoic acid a N,

Semantic acid CnKeto acid COOH 3 10: 70-100 0. OH; H eno-Blllanlc acid.

E=O --s 70C. N & On Lactone 25- 7 oxosapogenin My invention may be further illustrated by the following examples:

Example 1 Dihydrosarsasapogenin, M. P. 165166 C., is prepared by the method described by Marker and Rohrmann, J. Am. Chem. Soc., 61, 846 (1939).

A mixture of 20 g. of dihydrosarsasapogenin, M. P. 165166 C., and 200 cc. of acetic anhydride is refluxed for thirty minutes. The acetic anhydride is evaporated in vacuo and the residual sirup dissolved in 400 cc. of acetic acid. The well stirred solution is heated at 90-95? C. on a steam-bath while 44 g. of chromic anhydride in 250 cc. of 80% acetic acid is added over a period of two hours, after which the mixture is heated for an additional two hours. The mixture is concentrated in vacuo to a volume of about 100 cc. The residual material is diluted with water and the precipitated solids taken up in ether. The

ethereal solution, after thorough washing with water, is washed twice with 3% sodium hydroxide solution to remove the acidic fraction.

The sodium hydroxide washings containing the acidic material from the oxidation is heated on the steam-bath for twenty minutes to complete the hydrolysis. The cooled mixture is acidified with hydrochloric acid and the precipitated acids taken up in ether. Upon standing, the ethereal solution deposits 600 mg. of small compact white crystals which are recrystallized once from methanol to givea product, M. P. 285-288 C. dec. This is the C22 keto acid, M. P. 285-287 C. dec.

This substance, 3-(p)-hydroxy-16-keto-bisnor-cholanic acid, may be represented by the formula:

Example 2 To a solution of ,1 g. of'sarsasapogenin acetate in cc. of aceticacid at 20 C. is added 50 cc. of l N aqueous potassium permanganate solution. The temperature is maintained at 18 to 20 C. for fifteen hours, after which the mixture is diluted with water and extracted with ether. The

tion with hydrochloric acid yields a white solid which is taken up in ether and crystallized from this solvent to give 225 mg. of white crystals. The

product is recrystallized from methanol to give a product, M. P. 286-288 C. dec. 7 This is the C22 keto acid identical .with that described'in Exl l- Example 3 Evaporation of the ether solution containing the neutral products gives a sirup which is hydrolyzed with hot ethanolic potassium hydroxide. Dilution of the resulting solution with water gives a slight precipitate which is taken up in ether and discarded. The aqueous alkaline solution is acidified with hydrochloric acid and the precipitated material taken up in ether. After sublimation in high vacuum at ISO-180 the product is crystallized from ether-pentane to give white needles, M. P. 198-200 C. This is the C22 lactone related to sarsasapogenin. It may be represented by the formula:

CH3 CH3 To a solution of 800 mg. of the above mentioned lactone in 30 cc. of glacial acetic acid is added a solution of 2 g. of chromic anhydride in 40 cc. of 80% acetic acid. The resulting solution is heated on the steam-bath at 90 C. for three hours. The mixture is diluted with water and extracted with ether. The ethereal extract is washed well with water and then extracted with 3% sodium hydroxide solution. The sodium hydroxide extract is heated for twenty minutes on the steam-bath, cooled, acidified with hydrochloric acid and the solid acid extracted with ether. The ether upon slow evaporation deposits compact white crystals. These, after crystallization from ether-methanol, melted at 285-287 C. dec., and give no depression with a sample of the C22 keto acid, M. P. 285-287 C- Treatment of the methyl ester of the keto acid with boiling acetic anhydride for thirty minutes followed by decomposition of the excess acetic anhydride with water yields a methyl ester acetate which crystallizes from ether-pentane as compact white crystals, M. P. l98-199.5 C.

Treatment of the acid with hydroxylamine hydrochloride under the usual conditions gives an oxime which crystallizes from aqueous methanol as small compact white crystals, M. P. 206-208 C. dec.

Example 4 To a solution of 20 g. of sarsasapogenin acetate in 500 cc. of glacial acetic acid at 60-70 is added slowly over a period of 4 hours, a solution of 12 g. of chromic anhydride in 200 cc. of 90% acetic acid. The mixture is stirred an hour longer, then alcohol is added and the solution evaporated to a sirup. This sirup is dissolved in ether and the ethereal solution extracted with 3% sodium hydroxide solution.

The alkaline extract is warmed On a steambath for a few minutes and then it is cooled and acidified. The precipitated solid is crystallized from dilute acetone to give sarsasapogenoic acid of melting point 187-189 C,

The mother liquor is evaporated to dryness and the residue dissolved in a small amount of ether.

After this concentrated ethereal solution has stood at room temperature for several days, small compact white crystals are deposited. These are collected, washed with ether and recrystallized from methanol to give the C22 keto acid derived from sarsasapogenin. It may be represented by the following formula:

ether, acetone, chloroform and ethyl acetate, but

it is fairly soluble in methanol and ethanol.

The methyl ester of the above C22 keto acid may be obtained by allowing a suspension of 200 mg. of the keto acid in 20 cc. of methanol-ether (1:1) containing an excess of diazomethane to stand over night. Then the solvent is evaporated and the residue crystallized from the ether-pentane to give clusters of small plates which melt at 124-126" C., solidify at 127 C. and remelt at 159 C.

Other esters of this acid may be prepared, for example, by treatment with an appropriate alcohol and a suitable catalyst. For example, 200 mg. of the C22 keto acid may be refluxed with cc. of 1% ethyl alcoholic hydrogen chloride for three hours. Then the solution is concentrated to a small volume, diluted with water and extracted with ether. The ethereal layer is separated, washed with dilute sodium hydroxide solution and water, and the ether evaporated. The residue is crystallized from ether-pentane to give white needles of the ethyl ester, melting point 163-464 C.

Derivatives of the ketone grouping may be obtained by treating the acid or its esters with ketone reagents having a reactive NHz grouping. For example, a solution of 100 mg. of the C22 keto acid, 100 mg. of semicarbazide hydrochloride and mg. of sodium acetate in 10 cc. of alcohol a d 2 cc. of water is refluxed on the steam-bath for one hour. The solution is diluted with Water and the white solid collected and crystallized from ether to give a semicarbazone of melting point 204-207 C. dec.

Other sapogenins may be treated in accordance with the directions of this example to give analogous products. Such sapogenins include tigogenin, gitogenin and diosgenin. The latter contains a nuclear double bond which is protected by addition of bromine prior to oxidation. Instead of protecting the hydroxyl groups by acetylation during the oxidation, other protecting agents may be used. For example, the sapogenin may be benzoylated, benzylated or halogenated, for example, by treatment with phosphorus pentachloride in carbon disulfide.

Example 5 To 10 cc. of fuming nitric acid maintained at 20 C. by an external cooling bath is added 1 g. of sarsasapogenin acetate in small portions. After awhile when the sarsasapogenin acetate has completely dissolved, the solution is diluted with water and the gummy precipitate collected. This gummy precipitate is boiled for a short time with alcoholic sodium hydroxide. Then the solution is diluted, extracted with ether and the alkaline layer separated. This alkaline extract is acidified and the precipitated acid taken up in ether. The ethereal layer is separated and most of the ether removed. On standing for several days, this concentrated ethereal solution deposits crystals of the C22 keto acid derived from sarsasapogenin. This acid has the formula C22H34O4, and it melts at 285-287 C. dec. (gas evolution).

It will be apparent that in View of this disclosure my invention is capable of numerous variations with regard to conditions of reaction, reagents and sapogenins employed.

Steroidal sapogenins on which this invention may be practiced include not only the aglycones of the naturally occurring steroidal sapogenins, but also their nuclear transformation products, 1. e., the substances derived from the aglycones by changes in rings A and/or B which leave the side chain attached to ring D still intact and like that in the aglycones. Thus, this invention may be practiced on steroidal sapogeninssuch as smilagenin, sarsasapogenone, 3-desoxysarsasapogenin, the sarsasapogenyl chlorides, and the like.

It is apparent that sapogenins having in the side chain either configuration with regard to C22 may be employed in practicing this invention, that is to say, compounds having a side chain either of the type of sarsasapogenin or of the type tigogenin may be oxidized in the manner set forth in this specification, and regardless of the configuration of the side chain of the sapogenin oxidized the structure of the oxidation products remain the same, for example, sarsasapogenin and isosarsasapogenin give the same oxidation products.

What I claim as my invention is:

1. Process for obtaining sapogenin oxidation products which comprises subjecting a member of the class consisting of steroidal sapogenins, sapogentic acids, dihydrosapogenins, sapogenoic acids, and C22 lactones, to vigorous oxidation at 40-100 C. by treatment with a strong oxidizing agent after previously protecting hydroxyl groups therein with a member of the class consisting of acylating agents, aralkylating agents, and halogenating agents, thereby producing a C22 keto acid having in ring D the structure separating the alkali soluble fraction, thereby ootaining an alkali metal salt of a steroid acid having in ring D the structure,

CH; (EH. 0

where M represents an alkali metal, and hydrolyzing said fraction.

2. Process for preparing 3-e-hydroxy-16-ketobis-nor-cholanic acid which comprises subjecting sarsasapogenin acetate to oxidation at 40-100 C. by treatment with a strong oxidizing agent, separating the alkali soluble fraction, hydrolyzing said fraction and crystallizing 3-,8-hydroxy-16- keto-bis-nor-cholanic acid.

3. An acid having the formula CH: CH: CH: 11 00011 4. An ester having the formula CHI on coo on 5. A compound having the formula,

. CH: CH: CH:-

n-coon where R is selected from the class consisting of hydrogen and alkyl radicals and R1 is selected from the class consisting of 0H and groups hydrolyzable to ---OH.

RUSSELL EARL MARER. 

